The plasma membrane proton pump (H+-ATPase) in plants and fungi converts chemical energy into electrical energy, and generates the protonmotive force used to transport solutes into and out of all cells. My laboratory has cloned twelve different genes encoding the ATPase in A. thaliana, and is using light and electron microscopy techniques to identify their tissue and intracellular localization. When standard immunofluoresces microscopy is not sufficient for immunolocalization of antigens we will use the confocal microscope at the IMR. We will high pressure freeze samples for better preservation of antigenicity. After freezing the samples will be freeze substituted, embedded, sectioned and immunogold labeled. Immunolabeling will be observed by TEM. For this purpose, we are using two antibody preparations ( I ) a standard approach, using antibody generated against proteins obtained via expression in E. coli, (2) generation of recombinant plants containing a foreign 'epitope' (either mammalian c-myc or bacterial glucuronidase) attached to the carboxy terminus of the ATPase polypeptide. Our present studies focus on an ATPase gene isoform which is expressed in phloem cells.